The long-term goal of this project is the study of the fundamental biological processes which control the inheritance, growth, and replication of mitochondria in mammalian cells. These studies should also provide information concerning the interrelationship between the nucleus and the mitochondria as well as answers to questions about the informational content of mitochondrial DNA. In the shorter term we hope to establish conditions and procedures for bringing about the transfer of mitochondria and/or mitochondrial genetic material from one cell to a cell of another species and for their establishment in the recipient cell with the production of a stable transformed clone. Specifically we will determine how many and which human chromosomes are required to maintain human mitochondria using interspecific cell fusions. Chloramphenicol (CAP)-resistant human cells will be fused to mouse cells under conditions where human chromosomes segregate and human mitochondria are selected for (plus CAP). The resulting hybrids will be examined by enzymatic, isozyme and karyotype analysis. They would also be examined for human, mouse and recombinant mitochondrial DNA. We will correlate the retention of human mtDNA and the retention of specific human chromosomes. Another approach will be to fuse enucleated CAP- resistant human cells with CAP-sensitive mouse cells and select for a CAP-resistant cybrid (the fusion product between an enucleated and a nucleated cell). Following the demonstration of cytoplasmic transfer and stable inheritance of CAP-resistance intraspecifically, it becomes obvious that other criteria would be necessary to establish cytoplasmic inheritance in mammalian cells. Three basic experiments will be undertaken to confirm mitochondrial inheritance: 1) "rescue" of cells treated with ethidium bromide by fusion with untreated cells or with untreated cytoplasts; 2) segregation of cytoplasmic genes independently of the nuclear genes and 3) transfer of CAP resistance to a sensitive cell either by isolated mitochondria or purified mitochondrial DNA.